DOCSLIB.ORG

Investigation of Multiple-Dynein Transport of Melanosomes by Non- Invasive Force Measurement Using Fluctuation Unit 

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Investigation of Multiple-Dynein Transport of Melanosomes by Non- Invasive Force Measurement Using Fluctuation Unit  Title: Investigation of multiple-dynein transport of melanosomes by non- invasive force measurement using fluctuation unit Shin Hasegawa1, Takashi Sagawa2, Kazuho Ikeda3, Yasushi Okada3,4,5, and Kumiko Hayashi1,* 1Department of Applied Physics, Graduate School of Engineering, Tohoku University, Sendai, Japan 2Advanced ICT Research Institute, National Institute of Information and Communications Technology, Kobe, Japan 3Laboratory for Cell Dynamics Observation, Center for Biosystems Dynamics Research, RIKEN, Osaka, Japan 4Department of Physics and Universal Biology Institute, Graduate School of Science, The University of Tokyo, Tokyo, Japan 5Department of Physics, Universal Biology Institute, and the International Research Center for Neurointelligence (WPI-IRCN), The University of Tokyo, Tokyo, Japan *Correspondence: [email protected] (K.H.) 1 Pigment organelles known as melanosomes disperse or aggregate in a melanophore in response to hormones. These movements are mediated by the microtubule motors kinesin-2 and cytoplasmic dynein. However, the force generation mechanism of dynein, unlike that of kinesin, is not well understood. In this study, to address this issue, we investigated the dynein-mediated aggregation of melanosomes in zebrafish melanophores. We applied the fluctuation theorem of non-equilibrium statistical mechanics to estimate forces acting on melanosomes during transport by dynein, given that the energy of a system is related to its fluctuation. Our results demonstrate that multiple force-producing units cooperatively transport a single melanosome. Since the force is generated by dynein, this suggests that multiple dyneins carry a single melanosome. Cooperative transport has been reported for other organelles; thus, multiple- motor transport may be a universal mechanism for moving organelles within the cell. 2 Introduction Cellular cargo is transported through the microtubule network of eukaryotic cells by motor proteins1,2. This active transport system delivers materials more rapidly than the passive transport by diffusion occurring in prokaryotic cells. The importance of cargo transport is underscored by the fact that defects in this process in neurons are associated with neuronal diseases such as Alzheimer’s, Parkinson’s, and Huntington’s disease3,4. Multiple-carrier (motor) transport of cargo has recently been proposed5-13 as a mechanism for maintaining the stability of the intracellular transport system by increasing physical quantities such as force and run length. Since the cytosol has a higher viscosity than water14-16, a significant amount of force must be generated through cooperation of multiple motors to overcome friction and haul cargo in cells. In order to investigate force generated by motors acting on a single cargo, a non- invasive force measurement11-13 was recently developed based on the fluctuation theorem of non-equilibrium statistical mechanics17-22. Fluctuation in small systems is not merely due to random motion, but is related to system energetics through physical theorems. The position of a moving cargo can be obtained non-invasively by fluorescence microscopic observation of cells, and the fluctuation of its position is easily observed with high time resolution. Using this fluctuating motion, a fluctuation unit (χ) constructed based on the fluctuation theorem is calculated as a force indicator11-13. The physical background of χ was explained and discussed in the reference13. The parameter χ was previously measured for the transport of synaptic vesicle precursors (SVPs) by the kinesin superfamily protein UNC-104 in the DA9 neuron of Caenorhabditis elegans12. In wild-type worms, the distribution of χ was spread over several clusters, implying the existence of several force-producing units (FPUs); this in turn indicates that a single SVP was carried by multiple UNC-104 motors, which are the generators of force. Measurements of χ revealed that mutant worms lacking ARL-8—an SVP-bound Arf-like small guanosine triphosphatase that relieves the autoinhibition of the motor, which is critical for avoiding unnecessary consumption of ATP when the motor is not bound to an SVP23—had fewer FPUs than their wild-type counterparts12. In this study, we investigated χ for the transport of melanosomes, organelles filled with the melanin pigment, in zebrafish melanophores. Melanosomes are transported by microtubule motors, kinesin-2 and cytoplasmic dynein, and the actin motor myosin-V24. They disperse or aggregate in response to hormones in melanophores. The primary physiological purpose for movement of melanosomes in animals is colour change. Particularly, we focused 3 on the aggregation process of melanosomes transported by dynein. There are several advantages of measurement of χ in this transport system. Firstly, because melanin pigment is black and easily observed by bright-field microscopy, the recording rate can be increased up to 800 frames per second (fps). Such a high-speed recording enables accurate measurement of fluctuation in the position of melanosomes. Secondly, we can decrease the number of dynein motors using the inhibitor ciliobrevin, which was recently identified25 and investigated for many biological phenomena related to dynein25-29, thereby allowing observation of the behaviour of fluctuation unit χ in response to the decrease in motors. Thirdly, the force generation mechanism of dynein is still controversial and therefore worth studying. The stall force value for a single dynein molecule was reported to be 1 pN9,10,30, though a few studies have reported values of 5–10 pN31-33. Fourthly, the behaviour of χ remains to be evaluated in non-neuronal cells. When considered together, melanosome transport is a suitable system to explore χ. In our experiments, the quantal behaviour of χ was observed in melanosome transport by dynein. We concluded that several FPUs carry a single melanosome similarly to cargo transport in neurons11-13. Further, we found that the number of FPUs measured by χ was dramatically decreased by the addition of ciliobrevin, as expected. We anticipate that the non- invasive force measurement using the fluctuation unit (χ) will be applied to a wide range of cargo transport in eukaryotic cells to elucidate the physical mechanisms in which some human diseases are deeply rooted. 4 Results Observation of melanosome transport Melanophores removed from zebrafish scales by enzymatic treatment were cultured in a glass- bottom dish. After 1 day of culture, the motion of melanosomes was observed by bright-field microscopy. Following addition of hormone (epinephrine) to the dish, melanosomes aggregated to the centre of a melanophore. Because the minus ends of microtubules are attached to the centrosome, the motion towards the nucleus is caused by the action of dynein. A schematic of this transport is represented in Fig. 1. Although they were crowded at the start of migration, individual melanosomes could be tracked after several tens of seconds, when nearly all melanosomes were aggregated. The direction of movement was set as the plus X direction (e.g. red line in Fig. 2a). Except for melanosomes close to the nucleus where cells had greater thickness, we considered that the motion in the Z direction was less than that in the X-Y plane since the images of melanosomes were focused during the runs. The centre position (X) of the melanosome was calculated from the recorded images, and the time course of X was obtained (Fig. 2b). Directional movement was defined as movement at a velocity greater than 100 nm/s, as in a previous study on pigment granule transport5. The fluctuation in melanosome position during directional transport, which was observed at a high recording rate of 800 fps (inset in Fig. 2b), was mainly due to thermal noise, stochastic stepping of motors accompanied by ATP hydrolysis, and collision of melanosomes with other organelles and cytoskeletons. Calculation of for melanin pigment transport The constant velocity segment (red area in Fig. 2b) of the position (X) of a melanosome lasting 0.5–1 s was identified by visual inspection for calculation of the fluctuation unit χ (equation (4)). The effect of this selection on the calculation of χ was later evaluated by the bootstrapping method. Based on this segment, we calculated X = X(t + t) − X(t) (e.g. inset in Fig. 2b) and its probability distribution (P(X)) (Fig. 2c for the case t = 37.5 ms), where the value of t ranged from 1.25 to 37.5 ms. P(X) was well fitted by a Gaussian function (solid black curve in Fig. 2c). Because X < 0 could not be observed for a large t because of the limitations of our experiment, P(X) for X > 0 was fitted with a Gaussian function to calculate the ratio P(X)/P(–X) in equation (4). The Gaussian function (equation (5)) was used to fit P(X) after evaluating the Brownian noise power spectrum density S(f) (equation (11)) of X for small f (Supplementary Fig. S1). defined in equation (4) was calculated using equation (6) for each t (Fig. 2d); as a 5 function of t converged to a constant value (*) after relaxation ( 20 ms), which is attributable to the microscopic environment around the melanosome as well as the enzymatic cycle time13. Note that from a physical point of view, the convergence implied that the macroscopic motion of X whose time scale is about 20 ms obeys the simple equation (3). Then, equation (7) was considered for the constant value *. The error of * according to range selection of the constant velocity segment was estimated to be 15% based on the principle of bootstrapping. We first selected 10 different partial segments from the original constant velocity segments; was then calculated for each partial segment (thin curves in Fig. 2d). It should be noted that the length of partial segments was half that of the original segment. Melanosome transport by multiple dynein motors The procedures carried out in Fig. 2b–d for a single melanosome were repeated for 62 different melanosomes from two different melanophore preparations. The calculated values of for the 62 melanosomes are shown in Fig. 3a.
Load more

Recommended publications
  • Tracking Melanosomes Inside a Cell to Study Molecular Motors and Their Interaction
    Tracking melanosomes inside a cell to study molecular motors and their interaction Comert Kural*, Anna S. Serpinskaya†, Ying-Hao Chou†, Robert D. Goldman†, Vladimir I. Gelfand†‡, and Paul R. Selvin*§¶ *Center for Biophysics and Computational Biology and §Department of Physics, University of Illinois at Urbana–Champaign, Urbana, IL 61801; and †Department of Cell and Molecular Biology, Northwestern University School of Medicine, Chicago, IL 60611 Communicated by Gordon A. Baym, University of Illinois at Urbana–Champaign, Urbana, IL, January 9, 2007 (received for review June 4, 2006) Cells known as melanophores contain melanosomes, which are membrane organelles filled with melanin, a dark, nonfluorescent pigment. Melanophores aggregate or disperse their melanosomes when the host needs to change its color in response to the environment (e.g., camouflage or social interactions). Melanosome transport in cultured Xenopus melanophores is mediated by my- osin V, heterotrimeric kinesin-2, and cytoplasmic dynein. Here, we describe a technique for tracking individual motors of each type, both individually and in their interaction, with high spatial (Ϸ2 nm) and temporal (Ϸ1 msec) localization accuracy. This method enabled us to observe (i) stepwise movement of kinesin-2 with an average step size of 8 nm; (ii) smoother melanosome transport (with fewer pauses), in the absence of intermediate filaments (IFs); and (iii) motors of actin filaments and microtubules working on the same cargo nearly simultaneously, indicating that a diffusive step is not needed between the two systems of transport. In concert with our previous report, our results also show that dynein-driven retro- grade movement occurs in 8-nm steps. Furthermore, previous studies have shown that melanosomes carried by myosin V move 35 nm in a stepwise fashion in which the step rise-times can be as long as 80 msec.
    [Show full text]
  • Role of Cdc42 in Melanosome Transfer 1443 Approximate Ratio of 1:1 in KGM
    Research Article 1441 Filopodia are conduits for melanosome transfer to keratinocytes Glynis Scott, Sonya Leopardi, Stacey Printup and Brian C. Madden Department of Dermatology, University of Rochester School of Medicine and Dentistry, Rochester, NY, USA Author for correspondence (e-mail: [email protected]) Accepted 4 January 2002 Journal of Cell Science 115, 1441-1451 (2002) © The Company of Biologists Ltd Summary Melanosomes are specialized melanin-synthesizing cultured with keratinocytes induced a highly dendritic organelles critical for photoprotection in the skin. phenotype with extensive contacts between melanocytes Melanosome transfer to keratinocytes, which involves and keratinocytes through filopodia, many of which whole organelle donation to another cell, is a unique contained melanosomes. These results suggest a unique role biological process and is poorly understood. Time-lapse for filopodia in organelle transport and, in combination digital movies and electron microscopy show that filopodia with our previous work showing the presence of SNARE from melanocyte dendrites serve as conduits for proteins and rab3a on melanosomes, suggest a novel model melanosome transfer to keratinocytes. Cdc42, a small system for melanosome transfer to keratinocytes. GTP-binding protein, is known to mediate filopodia formation. Melanosome-enriched fractions isolated from Movies available on-line human melanocytes expressed the Cdc42 effector proteins PAK1 and N-WASP by western blotting. Expression of Key words: Melanosome, Melanocyte, Cdc42, Filopodia, constitutively active Cdc42 (Cdc42V12) in melanocytes co- Keratinocyte Introduction microscopy of cultured cells, which allowed direct Melanosomes are organelles unique to melanocytes that visualization of melanosome movement and modifiers of function in the synthesis of melanin, a complex pigment actin, microtubules and their motor proteins.
    [Show full text]
  • Primary Cilia Membrane Assembly Is Initiated by Rab11 and Transport Protein Particle II (TRAPPII) Complex- Dependent Trafficking of Rabin8 to the Centrosome
    Primary cilia membrane assembly is initiated by Rab11 and transport protein particle II (TRAPPII) complex- dependent trafficking of Rabin8 to the centrosome Christopher J. Westlakea,1, Lisa M. Bayeb, Maxence V. Nachuryc, Kevin J. Wrighta, Karen E. Ervina, Lilian Phua, Cecile Chalounia, John S. Beckd,e, Donald S. Kirkpatricka, Diane C. Slusarskib, Val C. Sheffieldd, Richard H. Schellera,1, and Peter K. Jacksona,1 aGenentech, South San Francisco, CA 94080; Departments of bBiology and dPediatrics and eHoward Hughes Medical Institute, University of Iowa, Iowa City, IA 52242; and cDepartment of Molecular and Cellular Physiology, Stanford University, Stanford, CA 94305 Contributed by Richard H. Scheller, December 18, 2010 (sent for review August 19, 2010) Sensory and signaling pathways are exquisitely organized in pri- Rab8 ciliary localization (3, 6) and its presence in Golgi/trans- mary cilia. Bardet-Biedl syndrome (BBS) patients have compromised Golgi membranes (7) suggest that Rab8 could participate in ciliary cilia and signaling. BBS proteins form the BBSome, which binds membrane assembly. Previously we reported Rabin8 at the cen- Rabin8, a guanine nucleotide exchange factor (GEF) activating the trosome (3), a localization that probably is important for Rab8 fi Rab8 GTPase, required for ciliary assembly. We now describe serum- ciliary traf cking. A candidate Rab8 GTPase activating protein regulated upstream vesicular transport events leading to centrosomal (GAP) (XM_037557) has been described and prevents cilia for- Rab8 activation and ciliary membrane formation. Using live micros- mation when overexpressed in cells; moreover, a catalytically in- copyimaging,we showthatupon serum withdrawalRab8 is observed active mutant localized to the cilia (6). An emerging mechanism for organizing vesicular transport to assemble the ciliary membrane in ∼100 min.
    [Show full text]
  • Vimentin Intermediate Filaments in Fish Melanophores
    Vimentin intermediate filaments in fish melanophores F. K. GYOEVA Institute of I'mlein Research, Academy of Scienc of the USSR, 142292 I'ushchino, Moscmv Region, USSR E. V. LEONOVA, V. I. RODIONOV and V. I. GELFAND* A. N. Belozersky Laboratory of Molecular Biology and Bioorganic Chemistry, Moscmv State University, 119S99 Moscmv, USSR * Author for correspondence Summary The distribution and chemical composition of as has been found in other cell types. Trans- intermediate filaments in cultured melanophores mission electron microscopy confirmed the pres- of two teleost species - Gymnocorymbus ternetzi ence of intermediate filaments in melanophores. and Pterophyllum scalare - were studied by im- Immunoblotting experiments showed the pres- munofluorescence staining and immunoblotting ence of the intermediate filament protein vimen- techniques. The immunofluorescence staining of tin in melanophore lysates. Therefore, teleost the melanophores with monoclonal and poly- melanophores possess a developed radial system clonal antibodies to the intermediate filament of vimentin intermediate filaments. protein vimentin revealed a system of fibrils radiating from the cell centre. These fibrils were Key words: melanophore, intermediate filaments, resistant to 0-6M-KC1 and nocodazole treatments vimentin. Introduction be involved. Microtubules form a well-developed radial pattern in fish melanophores and their disruption Melanophores are highly specialized cells, containing a inhibits pigment granule movement (Schliwa, 1981; lot of pigment granules known as melanosomes. Stearns, 1984). In contrast to microtubules, the system Teleost melanophores can aggregate melanosomes to of actin microfilaments in melanophores is poorly the cell centre or disperse them throughout the cyto- developed. Sparse microfilaments have been found in plasm. These melanosome movements, which deter- the cell cortex and in the cell surface microvilli mine the colour changes of animals, are governed by (Schliwa et al.
    [Show full text]
  • Myosin Cooperates with Microtubule Motors During Organelle Transport in Melanophores Stephen L
    Brief Communication 161 Myosin cooperates with microtubule motors during organelle transport in melanophores Stephen L. Rogers and Vladimir I. Gelfand Melanophores offer an outstanding system for the movement, several studies have pointed to the activity of study of intracellular motility. These cells aggregate a non-microtubule-based component as well [5–7]. These their pigment-filled melanosomes to the cell center or observations, together with the fact that pigment granules disperse them throughout the cytoplasm in response to in mammalian melanocytes are transported centrifugally hormonal modulation of intracellular cyclic AMP levels by a myosin [8,9], suggest that an actin-based transport in order to effect color changes in lower vertebrates [1]. mechanism may be a ubiquitous feature of melanosome Previous work from our laboratory demonstrated a role transport. This concept is also lent credence by the results for microtubule-based motors in melanosome transport of Rodionov et al., who demonstrate an association of and we succeeded in reconstituting their regulated melanosomes with actin filaments in fish melanophores motility along microtubules in vitro [2,3]. Here we [4]. To test directly the hypothesis that actomyosin- demonstrate that, in addition to microtubule-mediated driven motility contributes to melanosome transport, we motility, melanosomes purified from Xenopus examined the ability of these organelles to move along melanophores exhibit unidirectional movement along actin filaments in vitro. Using the Nitella myosin motility actin filaments in vitro as well. Immunoblotting analysis assay [10], we found that melanosomes purified from shows that these organelles possess the actin-based Xenopus melanophores attached to actin bundles, individ- organelle motor, myosin-V.
    [Show full text]
  • Role of Cytoplasmic Dynein in Melanosome Transport in Human Melanocytes
    Role of Cytoplasmic Dynein in Melanosome Transport in Human Melanocytes H. Randolph Byers, Mina Yaar, Mark S. Eller, Nicole L. Jalbert, and Barbara A. Gilchrest Department of Dermatology, Boston University School of Medicine, Boston, Massachusetts, U.S.A. Cytoplasmic dynein is a microtubule-associated transport of melanosomes from the cell body into the retrograde-directed motor molecule for transport of dendrites, whereas melanosome distribution was not membrane-bound organelles. To determine whether affected in sense-treated melanocytes. To determine cytoplasmic dynein is expressed in melanocytes, we whether ultraviolet irradiation modi®es cytoplasmic performed reverse transcriptase polymerase chain dynein expression, melanocyte cultures were reaction using melanocyte cDNA and primers com- exposed to increasing doses of solar-simulated irra- plementary to human brain cytoplasmic dynein diation, equivalent to a mild to moderate sunburn heavy chain. A polymerase chain reaction product of exposure for intact skin. Within 24 h, doses of 5 and the expected molecular size was generated and the 10 mJ per cm2 induced cytoplasmic dynein protein, identity was con®rmed by sequence analysis. whereas doses of 30 mJ per cm2 or more were asso- Western blotting of total melanocyte proteins reacted ciated with decreased levels of cytoplasmic dynein with an anti-intermediate chain cytoplasmic dynein compared with sham-irradiated controls. Our data antibody identi®ed the appropriate 74 kDa band. To show that cytoplasmic dynein participates in retro- determine whether cytoplasmic dynein plays a role grade melanosomal transport in human melanocytes in melanosome transport, duplicate cultures were and suggest that the altered melanosomal distribu- treated with cytoplasmic dynein antisense or sense tion in skin after sun exposure is due, at least in (control) oligodeoxynucleotides and the cells were part, to decreased cytoplasmic dynein levels result- observed by high-resolution time-lapse microscopy, ing in augmented anterograde transport.
    [Show full text]
  • Electron Tomography of Early Melanosomes: Implications for Melanogenesis and the Generation of Fibrillar Amyloid Sheets
    Electron tomography of early melanosomes: Implications for melanogenesis and the generation of fibrillar amyloid sheets Ilse Hurbaina,b, Willie J. C. Geertsc, Thomas Boudierd, Sergio Marcoe,f, Arie J. Verkleijc, Michael S. Marksg, and Grac¸a Raposoa,b,1 aInstitut Curie, Centre de Recherche, and bUnite´Mixte de Recherche 144, Centre National de la Recherche Scientifique, F-75248 Paris, France; cDepartment of Cellular Architecture and Dynamics, Universiteit Utrecht, Padualaan 8, 3584 CH, Utrecht, The Netherlands; dCentre National de la Recherche Scientifique Universite´Pierre et Marie Curie, Unite´Mixte de Recherche 7101, 75252 Paris Cedex 05, France; eInstitut Curie, Centre de Recherche, F-91405 Orsay, France; fInstitut National de la Sante´et de la Recherche Me´dicale, Unite´Mixte de Recherche 759, F-91405 Orsay, France; and gDepartment of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, PA, 19104-6100 Edited by David D. Sabatini, New York University School of Medicine, New York, NY, and approved August 27, 2008 (received for review April 15, 2008) Melanosomes are lysosome-related organelles (LROs) in which mel- appear to be composed largely of luminal fragments of Pmel17/ anins are synthesized and stored. Early stage melanosomes are gp100, a pigment-cell-specific integral membrane protein (11–14). characterized morphologically by intralumenal fibrils upon which Expression of Pmel17 alone in nonmelanocytic HeLa cells induces melanins are deposited in later stages. The integral membrane pro- the formation of similar fibrils within late endosomes (11), and a tein Pmel17 is a component of the fibrils, can nucleate fibril formation recombinant luminal Pmel17 M␣ fragment forms amyloid-like in the absence of other pigment cell-specific proteins, and forms fibrils upon renaturation in vitro (6).
    [Show full text]
  • Powering Membrane Traffic in Endocytosis and Recycling
    REVIEWS Powering membrane traffic in endocytosis and recycling Thierry Soldati* and Manfred Schliwa‡ Abstract | Early in evolution, the diversification of membrane-bound compartments that characterize eukaryotic cells was accompanied by the elaboration of molecular machineries that mediate intercompartmental communication and deliver materials to specific destinations. Molecular motors that move on tracks of actin filaments or microtubules mediate the movement of organelles and transport between compartments. The subjects of this review are the motors that power the transport steps along the endocytic and recycling pathways, their modes of attachment to cargo and their regulation. Endocytosis The emergence of eukaryotic cells was accompanied by attempt to consider details of the secretory pathway nor A plasma-membrane- the development of endomembrane systems that com- trafficking related to other organelles such as mitochon- associated process in which a partmentalize biochemical pathways and biosynthetic dria, chloroplasts or peroxisomes. We follow the journey eukaryotic cell engulfs processes. An evolutionary trend towards increasing of endocytic compartments on their inbound route from extracellular fluid or particles. complexity and subcellular specialization necessitated the plasma membrane to their centrally located destina- Phagocytosis the development of machineries for intercompartmental tions and then accompany recycling compartments back A form of endocytosis in which communication. Molecular assemblies evolved to con- to the
    [Show full text]
  • Melanin Transfer in the Epidermis: the Pursuit of Skin Pigmentation Control Mechanisms
    International Journal of Molecular Sciences Review Melanin Transfer in the Epidermis: The Pursuit of Skin Pigmentation Control Mechanisms Hugo Moreiras † , Miguel C. Seabra and Duarte C. Barral * iNOVA4Health, CEDOC, NOVA Medical School, NMS, Universidade NOVA de Lisboa, 1169-056 Lisboa, Portugal; [email protected] (H.M.); [email protected] (M.C.S.) * Correspondence: [email protected]; Tel.: +351-218-803-102 † Present address: The Charles Institute of Dermatology, School of Medicine, University College Dublin, D04 V1W8 Dublin, Ireland. Abstract: The mechanisms by which the pigment melanin is transferred from melanocytes and processed within keratinocytes to achieve skin pigmentation remain ill-characterized. Nevertheless, several models have emerged in the past decades to explain the transfer process. Here, we review the proposed models for melanin transfer in the skin epidermis, the available evidence supporting each one, and the recent observations in favor of the exo/phagocytosis and shed vesicles models. In order to reconcile the transfer models, we propose that different mechanisms could co-exist to sustain skin pigmentation under different conditions. We also discuss the limited knowledge about melanin processing within keratinocytes. Finally, we pinpoint new questions that ought to be addressed to solve the long-lasting quest for the understanding of how basal skin pigmentation is controlled. This knowledge will allow the emergence of new strategies to treat pigmentary disorders that cause a significant socio-economic burden to patients and healthcare systems worldwide and could also have relevant cosmetic applications. Citation: Moreiras, H.; Seabra, M.C.; Keywords: melanin; melanosome; melanocore; melanocyte; keratinocyte; skin pigmentation; Barral, D.C.
    [Show full text]
  • Autophagy and Cell Growth – the Yin and Yang of Nutrient Responses
    Commentary 2359 Autophagy and cell growth – the yin and yang of nutrient responses Thomas P. Neufeld Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, MN 55455, USA [email protected] Journal of Cell Science 125, 2359–2368 ß 2012. Published by The Company of Biologists Ltd doi: 10.1242/jcs.103333 Summary As a response to nutrient deprivation and other cell stresses, autophagy is often induced in the context of reduced or arrested cell growth. A plethora of signaling molecules and pathways have been shown to have opposing effects on cell growth and autophagy, and results of recent functional screens on a genomic scale support the idea that these processes might represent mutually exclusive cell fates. Understanding the ways in which autophagy and cell growth relate to one another is becoming increasingly important, as new roles for autophagy in tumorigenesis and other growth-related phenomena are uncovered. This Commentary highlights recent findings that link autophagy and cell growth, and explores the mechanisms underlying these connections and their implications for cell physiology and survival. Autophagy and cell growth can inhibit one another through a variety of direct and indirect mechanisms, and can be independently regulated by common signaling pathways. The central role of the mammalian target of rapamycin (mTOR) pathway in regulating both autophagy and cell growth exemplifies one such mechanism. In addition, mTOR-independent signaling and other more direct connections between autophagy and cell growth will also be discussed. This article is part of a Minifocus on Autophagy. For further reading, please see related articles: ‘Ubiquitin-like proteins and autophagy at a glance’ by Tomer Shpilka et al.
    [Show full text]
  • Antioxidants
    antioxidants Article The Effect of Antioxidants on Photoreactivity and Phototoxic Potential of RPE Melanolipofuscin Granules from Human Donors of Different Age Magdalena M. Olchawa 1 , Grzegorz M. Szewczyk 1 , Andrzej C. Zadlo 1 , Michal W. Sarna 1 , Dawid Wnuk 2 and Tadeusz J. Sarna 1,* 1 Department of Biophysics, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, 30-387 Kraków, Poland; [email protected] (M.M.O.); [email protected] (G.M.S.); [email protected] (A.C.Z.); [email protected] (M.W.S.) 2 Department of Cell Biology, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, 30-387 Kraków, Poland; [email protected] * Correspondence: [email protected]; Tel.: +48-12-664-64-27 Received: 28 August 2020; Accepted: 19 October 2020; Published: 26 October 2020 Abstract: One of the most prominent age-related changes of retinal pigment epithelium (RPE) is the accumulation of melanolipofuscin granules, which could contribute to oxidative stress in the retina. The purpose of this study was to determine the ability of melanolipofuscin granules from younger and older donors to photogenerate reactive oxygen species, and to examine if natural antioxidants could modify the phototoxic potential of this age pigment. Electron paramagnetic resonance (EPR) oximetry, EPR-spin trapping, and time-resolved detection of near-infrared phosphorescence were employed for measuring photogeneration of superoxide anion and singlet oxygen by melanolipofuscin isolated from younger and older human donors. Phototoxicity mediated by internalized melanolipofuscin granules with and without supplementation with zeaxanthin and α-tocopherol was analyzed in ARPE-19 cells by determining cell survival, oxidation of cellular proteins, organization of the cell cytoskeleton, and the cell specific phagocytic activity.
    [Show full text]
  • The Ocular Albinism Type 1 (OA1) GPCR Is Ubiquitinated and Its Traffic Requires Endosomal Sorting Complex Responsible for Transp
    The ocular albinism type 1 (OA1) GPCR is ubiquitinated and its traffic requires endosomal sorting complex responsible for transport (ESCRT) function Francesca Giordanoa,b,c,1, Sabrina Simoesa,b,c, and Graça Raposoa,b,c,2 aInstitut Curie, Centre de Recherche, Paris F-75248, France; bStructure and Membrane Compartments, Centre National de la Recherche Scientifique, UMR144, Paris F-75248, France; and cCell and Tissue Imaging Facility, Infrastructures en Biologie Sante et Agronomie, Paris F-75248, France Edited by David D. Sabatini, New York University School of Medicine, New York, NY, and approved June 7, 2011 (received for review March 2, 2011) The function of signaling receptors is tightly controlled by their tion and function remain undefined. Posttranslational modification intracellular trafficking. One major regulatory mechanism within the by ubiquitination controls intracellular trafficking events within fi endo-lysosomal system required for receptor localization and down- the endo-lysosomal system (10, 11). Ubiquitin-modi ed mem- regulation is protein modification by ubiquitination and down- brane proteins delivered from the endocytic or biosynthetic stream interactions with the endosomal sorting complex responsible pathways can be recognized by components of the endosomal sorting complex responsible for transport (ESCRT) machinery for transport (ESCRT) machinery. Whether and how these mecha- for targeting to intraluminal vesicles (ILVs) of multivesicular nisms operate to regulate endosomal sorting of mammalian G bodies (MVBs) and for subsequent
    [Show full text]